Abstract Harnessing optogenetics to diagnose and therapeutically rewire cancer cell signaling In recent years, advances in microscopy, sequencing, and proteomics have had a dramatic impact on our ability to probe the inner workings of cells and tissues. These tools have revealed that cells are remarkably complex information processing devices: external inputs activate many intracellular networks with complex dynamics. Yet despite the progress in measuring the cell's outputs, we have lacked analogous control over the inputs we can deliver to probe and perturb these complex networks. By replacing chemical stimuli with light, optogenetic tools can be used to apply and remove inputs with high spatial and temporal precision. Here, I propose to bring this precise input control to bear on two long-standing problems in cancer biology, a field where it has so far had limited impact. First, I will develop an approach termed ?optogenetic profiling?, which aims to directly measure how growth signaling is altered in tumor cells by measuring cellular responses to a rich set of input stimuli. Rooted in engineering, this approach is akin to probing an electronic circuit with a different signals to characterize its function, and can be highly informative even when the exact wiring diagram is unknown. This approach may offer a key to interpreting genomic data, allowing us to group cell lines with different mutations into shared functional classes. It could also have a large impact on treatment: identifying which pathway is deregulated may immediately suggest which targeted pathway inhibitors will be effective. Second, I will explore how light-induced protein aggregation can be used to therapeutically rewire cancer cell signaling. Tumor cells rely on signaling changes that both amplify proliferation and suppress apoptosis, but current therapies are typically limited to inhibiting pro-growth signaling. Here, I will test whether light-induced co-clustering of signaling proteins can perform two other therapeutic functions: amplifying apoptotic signaling or diverting growth inputs to cell-death outputs. The studies proposed will not only uncover fundamental principles in cell signaling but could usher in new approaches for cancer diagnosis (by functionally profiling signaling pathway responses) and treatment (by engineering gain-of-fuction therapies based on signaling enzyme clustering).